Process for preparing glycopeptide phosphonate derivatives

ABSTRACT

Disclosed are processes for preparing glycopeptide phosphonate derivatives having an amino-containing side chain. Several of the process steps are conducted in a single reaction vessel without isolation of intermediate reaction products, thereby generating less waste and improving the overall efficiency and yield of the process.

CROSS-REFERENCE TO RELATED APPLICATIONS

This application is a continuation of U.S. application Ser. No.10/926,148, filed Aug. 25, 2004; which application is a continuation ofU.S. application Ser. No. 10/226,428, filed Aug. 23, 2002 (now U.S. Pat.No. 6,979,723); which application claims the benefit of U.S. ProvisionalApplication No. 60/314,831, filed on Aug. 24, 2001; the entiredisclosures of which are incorporated herein by reference in theirentirety.

BACKGROUND OF THE INVENTION

1. Field of the Invention

This invention is directed to novel processes for preparing derivativesof glycopeptide antibiotics. More specifically, this invention isdirected to multi-step processes for preparing phosphonate derivativesof glycopeptide antibiotics having an amino-containing side chain, thefirst two steps being conducted in a single reaction vessel withoutisolation of the intermediate reaction products.

2. Background

Glycopeptides (e.g. dalbaheptides) are a well-known class of antibioticsproduced by various microorganisms (see Glycopeptide Antibiotics, editedby R. Nagarajan, Marcel Dekker, Inc. New York (1994)). Many syntheticderivatives of such glycopeptides are also known in the art and thesederivatives are typically reported to have improved properties relativeto the naturally-occurring glycopeptides, including enhancedantibacterial activity. For example, U.S. patent application Ser. No.09/847,042, filed May 1, 2001, describes various glycopeptidephosphonate derivatives, some of which contain an amino-containing sidechain. Such phosphate derivatives are particularly useful as antibioticsfor treating gram-positive infections.

Accordingly, a need exists for new efficient processes which are usefulfor preparing phosphonate derivatives of glycopeptide antibiotics havingan amino-containing side chain.

SUMMARY OF THE INVENTION

The present invention provides novel processes for preparing phosphonatederivatives of glycopeptide antibiotics having an amino-containing sidechain. Among other advantages, the first two steps of the presentprocess are conducted in a single reaction vessel without isolation ofthe intermediate reaction products, thereby generating less waste andimproving the overall efficiency and yield of the process compared toprevious processes.

Specifically, in one of its aspects, this invention is directed to aprocess for preparing a compound of formula 1:

wherein

R¹ is selected from the group consisting of C₁₋₁₀ alkylene, C₂₋₁₀alkenylene and C₂₋₁₀ alkynylene;

R² is selected from the group consisting of C₁₋₂₀ alkyl, C₂₋₂₀ alkenyl,C₂₋₂₀ alkynyl, C₃₋₈ cycloalkyl, C₅₋₈ cycloalkenyl, C₆₋₁₀ aryl, C₂₋₉heteroaryl, C₂₋₉ heterocyclic, —R^(a)-Cy¹, —R^(a)—Ar¹—Ar²,—R^(a)—Ar¹—R^(b)—Ar², —R^(a)—Ar¹—O—R^(b)—Ar²;

R⁴ is C₁₋₁₀ alkylene;

R^(a) is selected from the group consisting of C₁₋₁₀ alkylene, C₁₋₁₀alkenylene and C₁₋₁₀ alkynylene;

R^(b) is selected from the group consisting of C₁₋₆ alkylene, C₁₋₆alkenylene and C₁₋₆ alkynylene;

Cy¹ is selected from the group consisting of C₃₋₈ cycloalkyl, C₅₋₈cycloalkenyl, C₆₋₁₀ aryl, C₂₋₉ heteroaryl, C₂₋₉ heterocyclic;

Ar¹ and Ar² are independently selected from C₆₋₁₀ aryl and C₂₋₉heteroaryl;

wherein each aryl, heteroaryl and heterocyclic group is optionallysubstituted with 1 to 3 substituents independently selected from thegroup consisting of C₁₋₆ alkyl, C₁₋₆ alkoxy, halo, hydroxy, nitro andtrifluoromethyl, and each heteroaryl and heterocyclic group containsfrom 1 to 3 heteroatoms selected from nitrogen, oxygen or sulfur;

or a salt thereof;

the process comprising:

(a) reacting vancomycin or a salt thereof, with a compound of formulaII:

wherein R¹ and R² are as defined herein; and R³ is a amine-labileprotecting group; and a reducing agent to form a compound of formulaIII:

wherein R¹, R² and R³ are as defined herein or a salt thereof;

(b) reacting the compound of formula III with an amine to provide acompound of formula IV:

wherein R¹ and R² are as defined herein, or a salt thereof; wherein step(a) and step (b) are conducted in the same reaction mixture withoutisolation of the intermediate from step (a); and

(c) reacting the compound of formula IV with formaldehyde and a compoundof formula V:

in the presence of a base to provide a compound of formula I, or a saltthereof.

In the above process, R¹ is preferably C₁₋₆ alkylene. More preferably,R¹ is C₁₋₂ alkylene. Still more preferably, R¹ is —CH₂—.

R² is preferably C₆₋₁₄ alkyl. More preferably, R² is C₈₋₁₂ alkyl. Stillmore preferably, R² is n-decyl.

In the process of this invention, R³ is an amino-protecting group whichis removed by treatment with an amine (i.e., a nucleophilic amine).Preferably, R³ is a group of formula (A):W—OC(O)—  (A)wherein W is selected from the group consisting of 9-fluorenylmethyl,3-indenylmethyl, benz[f]inden-3-ylmethyl,17-tetrabenzo[a,c,g,i]fluorenylmethyl,2,7-di-tert-butyl[9-(10,10-dioxo-10,10,10,10-tetrahydrothioxanthyl)]methyl,1,1-dioxobenzo[b]thiophene-2-ylmethyl, wherein the 9-fluorenylmethylgroup is optionally substituted with 1 to 3 substitutents selected fromthe group consisting of C₁₋₆ alkyl, halo, nitro and sulfo.

Preferably, W is 9-fluorenylmethyl, wherein the 9-fluorenylmethyl groupis optionally substituted with 1 to 3 substitutents selected from thegroup consisting of C₁₋₆ alkyl, halo, nitro and sulfo. More preferably,W is 9-fluorenylmethyl.

Preferably, R⁴ is C₁₋₆ alkylene. More preferably, R⁴ is C₁₋₄ alkylene.Still more preferably, R⁴ is —CH₂—.

In step (a), the reducing agent is preferably an amine/borane complex.More preferably, the reducing agent is pyridine/borane ortert-butylamine/borane; and still more preferably, the reducing agent istert-butylamine/borane.

In a preferred embodiment of this process, step (a) comprises the stepsof:

(i) combining vancomycin or a salt thereof with a compound of formula IIin the presence of base to form a reaction mixture;

(ii) acidifying the reaction mixture from step (i) with an acid; and

(iii) contacting the reaction mixture from step (ii) with a reducingagent.

In this preferred embodiment, the base in step (i) is preferably atertiary amine; more preferably, the base is diisopropylethylamine.

Preferably, the acid employed in step (ii) is trifluoroacetic acid oracetic acid.

In step (b), the amine employed is preferably ammonium hydroxide or aprimary amine. More preferably, the amine is ammonium hydroxide,methylamine or tert-butylamine; and still more preferably, the amine istert-butylamine.

In step (c), the base employed is preferably a tertiary amine.Preferably, the tertiary amine employed is diisopropylethylamine. In apreferred embodiment, the molar ratio of tertiary amine to compound offormula V is about 3:1 to about 5:1; more preferably, about 4:1.

DETAILED DESCRIPTION OF THE INVENTION

This invention relates to novel processes for preparing glycopeptidephosphonate derivatives having an amino-containing side chain. Whendescribing such processes, the following terms have the followingmeanings, unless otherwise indicated.

DEFINITIONS

The term “alkyl” refers to a monovalent saturated hydrocarbon groupwhich may be linear or branched. Unless otherwise defined, such alkylgroups typically contain from 1 to 20 carbon atoms. Representative alkylgroups include, by way of example, methyl, ethyl, n-propyl, isopropyl,n-butyl, sec-butyl, isobutyl, tert-butyl, n-pentyl, n-hexyl, n-heptyl,n-octyl, n-nonyl, n-decyl and the like.

The term “alkenyl” refers to a monovalent unsaturated hydrocarbon groupwhich may be linear or branched and which has at least one, andtypically 1, 2 or 3, carbon-carbon double bonds. Unless otherwisedefined, such alkenyl groups typically contain from 2 to 20 carbonatoms. Representative alkenyl groups include, by way of example,ethenyl, n-propenyl, isopropenyl, n-but-2-enyl, n-hex-3-enyl and thelike.

The term “alkynyl” refers to a monovalent unsaturated hydrocarbon groupwhich may be linear or branched and which has at least one, andtypically 1, 2 or 3, carbon-carbon triple bonds. Unless otherwisedefined, such alkynyl groups typically contain from 2 to 20 carbonatoms. Representative alkynyl groups include, by way of example,ethynyl, n-propynyl, n-but-2-ynyl, n-hex-3-ynyl and the like.

The term “alkylene” refers to a divalent saturated hydrocarbon groupwhich may be linear or branched. Unless otherwise defined, such alkylenegroups typically contain from 1 to 10 carbon atoms. Representativealkylene groups include, by way of example, methylene, ethane-1,2-diyl(“ethylene”), propane-1,2-diyl, propane-1,3-diyl, butane-1,4-diyl,pentane-1,5-diyl and the like.

The term “alkenylene” refers to a divalent unsaturated hydrocarbon groupwhich may be linear or branched and which has at least one, andtypically 1, 2 or 3, carbon-carbon double bonds. Unless otherwisedefined, such alkenylene groups typically contain from 2 to 10 carbonatoms. Representative alkenylene groups include, by way of example,ethene-1,2-diyl, prop-1-ene-1,2-diyl, prop-1-ene-1,3-diyl,but-2-ene-1,4-diyl, and the like.

The term “alkynylene” refers to a divalent unsaturated hydrocarbon groupwhich may be linear or branched and which has at least one, andtypically 1, 2 or 3, carbon-carbon triple bonds. Unless otherwisedefined, such alkynylene groups typically contain from 2 to 10 carbonatoms. Representative alkynylene groups include, by way of example,ethyne-1,2-diyl, prop-1-yne-1,2-diyl, prop-1-yne-1,3-diyl,but-2-yne-1,4diyl, and the like.

The term “alkoxy” refers to a group of the formula —O—R, where R isalkyl as defined herein. Representative alkoxy groups include, by way ofexample, methoxy, ethoxy, n-propoxy, isopropoxy, n-butoxy, sec-butoxy,isobutoxy, tert-butoxy and the like.

The term “aryl” refers to a monovalent aromatic hydrocarbon having asingle ring (i.e., phenyl) or fused rings (i.e., naphthalene). Unlessotherwise defined, such aryl groups typically contain from 6 to 10carbon ring atoms. Representative aryl groups include, by way ofexample, phenyl and naphthalene-1-yl, naphthalene-2-yl, and the like.

The term “cycloalkyl” refers to a monovalent saturated carbocyclichydrocarbon group. Unless otherwise defined, such cycloalkyl groupstypically contain from 3 to 10 carbon atoms. Representative cycloalkylgroups include, by way of example, cyclopropyl, cyclobutyl, cyclopentyl,cyclohexyl and the like.

The term “cycloalkenyl” refers to a monovalent unsaturated carbocyclichydrocarbon group having at least one carbon-carbon double bond in thecarbocyclic ring. Unless otherwise defined, such cycloalkenyl groupstypically contain from 5 to 10 carbon atoms. Representative cycloalkenylgroups include, by way of example, cyclopent-3-en-1-yl,cyclohex-1-en-1-yl and the like.

The term “halo” refers to fluoro, chloro, bromo and iodo; preferably,chloro, bromo and iodo.

The term “heteroaryl” refers to a monovalent aromatic group having asingle ring or two fused rings and containing in the ring at least oneheteroatom (typically 1 to 3 heteroatoms) selected from nitrogen, oxygenor sulfur. Unless otherwise defined, such heteroaryl groups typicallycontain from 5 to 10 total ring atoms. Representative heteroaryl groupsinclude, by way of example, monovalent species of pyrrole, imidazole,thiazole, oxazole, fur, thiophene, triazole, pyrazole, isoxazole,isothiazole, pyridine, pyrazine, pyridazine, pyrimidine, triazine,indole, benzofuran, benzothiophene, benzimidazole, benzthiazole,quinoline, isoquinoline, quinazoline, quinoxaline and the like, wherethe point of attachment is at any available carbon or nitrogen ringatom.

The term “heterocycle” or “heterocyclic” refers to a monovalentsaturated or unsaturated (non-aromatic) group having a single ring ormultiple condensed rings and containing in the ring at least oneheteroatom (typically 1 to 3 heteroatoms) selected from nitrogen, oxygenor sulfur. Unless otherwise defined, such heterocyclic groups typicallycontain from 2 to 9 total ring atoms. Representative heterocyclic groupsinclude, by way of example, monovalent species of pyrrolidine,imidazolidine, pyrazolidine, piperidine, 1,4-dioxane, morpholine,thiomorpholine, piperazine, 3-pyrroline and the like, where the point ofattachment is at any available carbon or nitrogen ring atom.

The term “vancomycin” is used herein in its art recognized manner torefer to the glycopeptide antibiotic known as vancomycin. See, forexample, R. Nagarajan, “Glycopeptide Anitibiotics”, Marcel Dekker, Inc.(1994) and references cited therein. The designation “N^(van)-” refersto substitution at the vancosamine nitrogen atom of vancomycin. Thisposition is also referred to as the N3″ position of vancomycin.Additionally, using a conventional vancomycin numbering system, thedesignation “29-” refers to the carbon atom position between the twohydroxyl groups on the phenyl ring of amino acid 7 (AA-7). This positionis also sometimes referred to as the “7d” or the “resorcinol position”of vancomycin.

The term “salt” when used in conjunction with a compound referred toherein refers to a salt of the compound derived from an inorganic ororganic base or from an inorganic or organic acid. Salts derived frominorganic bases include aluminum, ammonium, calcium, copper, ferric,ferrous, lithium, magnesium, manganic, manganous, potassium, sodium,zinc and the like. Particularly preferred are ammonium, calcium,magnesium, potassium and sodium salts. Salts derived from organic basesinclude salts of primary, secondary and tertiary amines, includingsubstituted amines, cyclic amines, naturally occuring amines and thelike, such as arginine, betaine, caffeine, choline,N,N′-dibenzylethylenediamine, diethylamine, 2-diethylaminoethanol,2-dimethylaminoethanol, ethanolamine, ethylenediamine,N-ethylmorpholine, N-ethylpiperidine, glucamine, glucosamine, histidine,hydrabamine, isopropylamine, lysine, methylglucamine, morpholine,piperazine, piperadine, polyamine resins, procaine, purines,theobromine, triethylamine, trimethylamine, tripropylamine, tromethamineand the like. Salts derived from acids include acetic, ascorbic,benzenesulfonic, benzoic, camphosulfonic, citric, ethanesulfonic,fumaric, gluconic, glucoronic, glutamic, hippuric, hydrobromic,hydrochloric, isethionic, lactic, lactobionic, maleic, malic, mandelic,methanesulfonic, mucic, naphthalenesulfonic, nicotinic, nitric, pamoic,pantothenic, phosphoric, succinic, sulfuric, tartaric, p-toluenesulfonicand the like. Particularly preferred are citric, hydrobromic,hydrochloric, maleic, phosphoric, sulfuric and tartaric acids.

The term “protecting group” or “blocking group” refers to a group which,when covalently attached to a function group such as an amino, hydroxyl,thiol, carboxyl, carbonyl and the like, prevents the functional groupfrom undergoing undesired reactions but which permits the function groupto be regenerated (i.e., deprotected or unblocked) upon treatment of theprotecting group with a suitable reagent. Representative protectinggroups are disclosed, for example, in T. W. Greene and P. G. M. Wuts,“Protective Groups in Organic Synthesis” 3rd Ed., 1999, John Wiley andSons, N.Y.

The term “amine-labile protecting group” refers to a protecting groupwhich is removed upon treatment with a suitable amine.

Process Conditions

The process of the present invention is conducted in three stepsbeginning with vancomycin or a salt thereof. The first step of theprocess is a reductive alkylation step which involves first combiningone equivalent of vancomycin or a salt thereof, with one or moreequivalents of an aldehyde of formula II:

wherein R¹, R² and R³ are as defined herein to form a imine and/orhemiaminal intermediate in situ.

The aldehydes of formula II employed in the process of the presentinvention are well-known in the art and are either commerciallyavailable or can be prepared by conventional procedures usingcommercially available starting materials and conventional reagents. Forexample, see WO 00/39156, published on Jul. 6, 2000, which describesvarious methods for preparing such aldehydes.

Typically, the vancomycin or a salt thereof and the aldehyde arecombined in an inert diluent in the presence of an excess amount of asuitable base to form a reaction mixture. Preferably, the inert diluentis N,N-dimethylformamide, N,N-dimethylacetamide, N-methylpyrrolidinone,acetonitrile/water, and the like or mixtures thereof Preferably, fromabout 1 to about 2 equivalents of the aldehyde are employed; morepreferably, about 1.1 to about 1.2 equivalents. In this reactionmixture, a mixture of imines and/or hemiaminals is believed to be formedbetween the aldehyde and the basic nitrogen atoms of vancomycin, i.e.,the vancosamine nitrogen atom and the N-terminal (leucinyl) nitrogenatom.

Formation of the imine and/or hemiaminal intermediate is typicallyconducted at a temperature ranging from about 0° C. to about 75° C.,preferably at ambient temperature (i.e., about 20-25° C.) for about 1 toabout 24 hours, preferably for about 6 to 12 hours, or until formationof the imine and/or hemiaminal is substantially complete.

Any suitable base may be employed to neutralize the vancomycin salt andto facilitate formation of the imine and/or hemiaminal, includingorganic bases, such as amines, alkali metal carboxylate salt (ire.,sodium acetate and the like) and inorganic bases, such as alkali metalcarbonates (i.e., lithium carbonate, potassium carbonate and the like).Preferably, the base is a tertiary amine including, by way ofillustration, triethylamine, diisopropylethylamine, N-methylmorpholine,and the like. A preferred base is diisopropylethylamine. The base istypically employed in a molar excess relative to vancomycin. Preferably,the base is used in an amount ranging from about 1.5 to about 3equivalents based on vancomycin; more preferably, about 1.8 to 2.2equivalents.

When formation of the imine and/or hemiaminal mixture is substantiallycomplete, the reaction mixture is acidified with an excess of acid. Anysuitable acid may be employed including, by way of illustration,carboxylic acids (e.g. acetic acid, trichloroacetic acid, citric acid,formic acid, trifluoroacetic acid, methanesulfonic acid, toluenesulfonicacid and the like), mineral acids (e.g. hydrochloric acid, sulfuricacid, or phosphoric acid), and the like. Preferably, the acid istrifluoroacetic acid or acetic acid. The acid is typically added in amolar excess relative to vancomycin (and the base). Preferably, the acidis used in an amount ranging from about 3 to about 6 equivalents basedon vancomycin; more preferably, about 3.5 to 5.5 equivalents.

While not wishing to be limited by theory, it is believed that the acidselectively hydrolyzes the imine and/or hemiaminal formed at theN-terminal amine of vancomycin in preference to the imine and/orhemiaminal formed at the vancosamine nitrogen atom. Acidification of thereaction mixture is typically conducted at a temperature ranging fromabout 0° C. to about 30° C., preferably at about 25° C., for about 0.25to about 2.0 hours, preferably for about 0.5 to about 1.5 hours.Preferably, a polar, protic solvent is added during this step including,by way of example, methanol, ethanol, propanol, isopropanol, butanol,ethylene glycol, and the like. Alternatively, a mixed polarprotic/non-protic solvent may be used, such as methanol/tetrahydrofuran,methanol/1,2-dimethoxyethane and the like

After acidification, the reaction mixture is contacted with a reducingagent to reduce the imine and/or hemiaminal. Any suitable reducing agentcan be employed which is compatible with the functionality present inthe glycopeptide. For example, suitable reducing agents include sodiumborohydride, sodium cyanoborohydride, zinc borohydride, sodiumtriacetoxyborohydride, pyridine/borane, tert-butylamine/borane,N-methylmorpholine/borane, ammonia/borane, dimethylamine/borane,triethylamine/borane, trimethylamine/borane, and the like. Preferredreducing agents are amine/borane complexes such as pyridine/borane andtert-butylamine/borane.

The reduction phase of the reaction is typically conducted at atemperature ranging from about 0° C. to about 30° C., preferably atabout 25° C., for about 0.5 to about 24 hours, preferably for about 1 toabout 6 hours, or until the reduction is substantially complete.Preferably, a polar, protic solvent is present during this reductionstep. The polar, protic solvent is preferably added during theacidification described above.

In contrast to prior procedures, the product of the reductive alkylationprocess is not isolated but the reaction mixture is contacted with anamine to remove the protecting group (i.e., R³) from the intermediateproduct. Any suitable amine may be used in this step of the process.Representative amines suitable for use include, by way of example,methylamine, ethylamine, tert-butylamine, triethylamine, piperidine,morpholine, ammonium hydroxide, 1,4-diazabicyclo[2.2.2]octane (DABCO)and the like. Preferred amines are methylamine, tert-butylamine,ammonium hydroxide and 1,4-diazabicyclo[2.2.2]octane.

This deprotection step is typically conducted at a temperature rangingfrom about 0° C. to about 60° C., preferably at about 40° C. to about45° C., for about 2 to about 60 hours, preferably for about 3 to about10 hours, or until the reaction is substantially complete. This step istypically conducted in an inert diluent, such as N,N-dimethylformamide,N,N-dimethylacetamide, N-methylpyrrolidinone, and the like. Theresulting compound of formula IV is readily isolated and purified byconventional procedures, such as precipitation, filtration and the like.

In the next step of the process, the compound of formula IV is contactedwith formaldehyde and a compound of formula V:

wherein R⁴ is as defined herein; in the presence of a base to provide acompound of formula I, or a salt thereof.

This step of the process is typically conducted by contacting oneequivalent of compound IV or a salt thereof with one or moreequivalents, preferably with about 2 to about 10 equivalents of acompound of formula V, and with an excess, preferably with about 4 toabout 5 equivalents, formaldehyde in the presence of a base.

Phosphonate compounds of formula V are either commercially available orcan be prepared by conventional procedures using commercially availablestarting materials and reagents. See for example, Advanced OrganicChemistry, Jerry March, 4th ed, 1992, John Wiley and Sons, New York,page 959; and Frank R. Hartley (ed.) The Chemistry of OrganophosphorousCompounds, vol. 1-4, John Wiley and Sons, New York (1996).Aminomethylphosphonic acid is commercially available from AldrichChemical Company, Milwaukee, Wisc.

The formaldehyde employed in this step of the process is typically addedin an aqueous solution, for example, as a 37 wt. % solution in wateroptionally containing about 5 to about 15 wt. % methanol (i.e.,Formalin).

Any suitable base may be used in this reaction including, for example,organic bases such as tertiary amines, and inorganic bases, such asalkali metal hydroxides (i.e., sodium hydroxide). Preferably, the baseis a tertiary amine including, by way of example, triethylamine,diisopropylethylamine, and the like. A preferred tertiary amine isdiisopropylethylamine. Preferably, the molar ratio of tertiary amine tocompound V is about 3:1 to about 5:1; more preferably, about 3.5:1 toabout 4.5:1; and still more preferably, about 4:1. Preferably, the pH ofthe reaction mixture is preferably about 10 to about 11.

Preferably, this reaction is conducted in an inert diluent, such aswater, acetonitrile/water and the like. In a preferred embodiment, thisstep of the process is conducted in acetonitrile/water or water havingv/v ratio ranging from about 3:2 to completely water.

This step of the process is typically conducted at a temperature rangingfrom about −20° C. to about 20° C., preferably at about −10° C. to about−5° C., for about 6 to about 48 hours, or until the reaction issubstantially complete.

The resulting compound of formula I or a salt thereof is isolated byconventional procedures including, precipitation, filtration and thelike. In a preferred isolation procedure, the pH of the reaction mixtureis adjusted to about 2 to about 3 by addition of a suitable acid, suchas aqueous hydrochloride acid. Preferably, the temperature of thereaction mixture is maintained below about 5° C. during acidification.Acetonitrile is then added to promote precipitation of the reactionproduct (i.e., a compound of formula I) and the resulting precipitate iscollected by filtration and optionally washed with additionalacetonitrile.

If desired, the reaction product can be further purified usingreverse-phase HPLC or other chromatographic methods. In a preferredembodiment, the product is purified using a resin as described inco-pending U.S. application Ser. No. 10/226,676, filed on Aug. 23, 2002;which application claims the benefit of U.S. Provisional Application No.60/314,712, filed on Aug. 24, 2001; the disclosures of which areincorporated herein by reference in their entirety.

Among other advantages, the process of the present invention providesfor improved yield, purity and selectivity, i.e., reductive alkylationat the vancosamine amino group is favored over reductive alkylation atthe N-terminus (e.g., the leucinyl group) by at least 10:1, morepreferably 20:1. Additionally, because the reductive alkylation anddeprotection steps are conducted in a single reaction vessel withoutisolation of the reaction intermediates, the process of the presentinvention is more efficient, provides a higher yield and generates lesswaste then previous processes.

The glycopeptide derivatives produced by the process of this inventionare useful as antibiotics. See, for example, U.S. patent applicationSer. No. 09/847,042, filed May 1, 2001; the disclosure of which isincorporated herein by reference in its entirety.

Additional details of the process of this invention are described in thefollowing Examples which are offered to illustrate this invention andare not to be construed in any way as limiting the scope of thisinvention.

EXAMPLES

In the examples below, the following abbreviations have the followingmeanings. Any abbreviations not defined have their generally acceptedmeaning. Unless otherwise stated, all temperatures are in degreesCelsius (° C.).

DIPEA=diisopropylethylamine

DMF=N,N-dimethylformamide

DMSO=dimethyl sulfoxide

eq.=equivalent

Fmoc=9-fluorenylmethoxycarbonyl

TFA=trifluoroacetic acid

In the following examples, vancomycin hydrochloride semi-hydrate waspurchased from Alpharma, Inc. Fort Lee, N.J. 07024 (Alpharma AS, OsloNorway). Other reagents and reactants are available from AldrichChemical Co., Milwaukee, Wisc. 53201.

Example A Preparation of N-Fmoc-Decylaminoacetaldehyde

Step A—Preparation of N-Fmoc-2-(n-Decylamino)ethanol

2-n-Decylamino)ethanol (2.3 g, 11 mmol, 1.1 eq) and DIPEA (2.0 mL, 11mmol, 1.1 eq) were dissolved in methylene chloride (15 mL) and cooled inan ice bath. 9-Fluorenylmethyl chloroformate (2.6 g, 10 mmol, 1.0 eq) inmethylene chloride (15 ml) was added, the mixture stirred for 30 minutesthen washed with 3 N hydrochloric acid (50 mL) twice and saturatedsodium bicarbonate (50 mL). The organics were dried over magnesiumsulfate, and the solvents removed under reduced pressure.N-Fmoc-2-(n-decylamino)ethanol (4.6 g, 11 mmol, 108%) was used withoutfurther purification.

Step B—Preparation of N-Fmoc-2-(n-Decylamino)acetaldehyde

To a solution of oxalyl chloride (12.24 mL) and methylene chloride (50mL) at −35 to −45° C. was added DMSO (14.75 g) in methylene chloride (25mL) over 20 minutes. The reaction mixture was stirred for 10 minutes at−35 to −45° C. A solution of N-Fmoc-2-(n-decylamino)ethanol (20.0 g) inmethylene chloride (70 mL) was added over 25 minutes and then stirred 40minutes at −35 to −45° C. Triethylamine (21.49 g) was then added and themixture stirred for 30 minutes at −10 to −20° C. The reaction mixturewas quenched with water (120 mL) followed by concentrated sulfuric acid(20.0 g) while maintaining the internal temperature at 0-5° C. Theorganic layer was isolated and washed with 2% sulfuric acid (100 mL)followed by water (2×100 mL). The organic solution was distilled undervacuum at 60° C. to about 100 mL. Heptane (100 mL) was added, thetemperature of the oil bath raised to 80° C. and the distillation wascontinued until the residual volume was 100 mL. More heptane (100 mL)was added and the distillation repeated to a volume of 100 mL. Theheating bath was replaced with a cold water bath at 15° C. The bath wascooled slowly to 5° C. over 20 minutes to start the precipitation of theproduct. The slurry was then cooled to −5 to −10° C. and the slurry wasstirred for 2 hours. The solid was then collected on a Buchner funneland washed with cold (−5° C.) heptane (2×15 mL). The wet solid was driedin vacuo to yield the title aldehyde.

Example 1 Preparation of N^(van)-2-(n-Decylamino)ethyl VancomycinHydrochloride

To a stirred mixture of 20 g (13.46 mmol) of vancomycin hydrochlorideand 6.526 g (15.48 mmol) of N-Fmoc-2-(n-decylamino)acetyldehyde wasadded 130 mL of N,N-dimethylformamide and 4.7 mL (26.92 mmol) ofN,N-diisopropylethylamine. The resulting mixture was stirred at roomtemperature under nitrogen for 15 hours, and 75 mL of methanol and 4.15mL of trifluoroacetic acid (53.84 mmol) were added at 0° C.successively. The mixture was stirred for 1 hour and 1.93 mL (15.48mmol) of borane-pyridine complex was added. The resulting mixture wasstirred for 4 hours at 0° C., and 80 mL (161.52 mmol) of a2 Mmethylamine in methanol was added. The resulting mixture was warmed toroom temperature and stirred for 50 hours, cooled to 0° C., and water(350 mL) was added dropwise. The mixture was acidified to pH 3.60 byslow addition of 11 mL of concentrated hydrochloric acid, andprecipitation occurred. The mixture was stirred for another 30 min andthen it was filtered through a Buchner funnel. The resulting wet cakewas washed with water (2×200 mL) and dried in vacuo for 16 hours to give9.8 g of crude N^(van)-2-(n-decylamino)ethyl vancomycin hydrochloride.This intermediate may then be used in step (c) of the process asdescribed in Example 3.

Example 2 Preparation of N^(van)-2-(n-Decylamino)ethyl VancomycinHydrochloride

To a 1L three-necked round bottom flask equipped with a mechanicalstirrer, a thermometer and a nitrogen bubbler was added 180 mL ofN,N-dimethylformamide (DMF). While stirring, 6.75 g (0.0160 mol) ofN-Fmoc-2-(n-decylamino)-acetyldehyde and 25 g (0.0168 mol) of vancomycinhydrochloride were added successively. The addition funnel was rinsedwith 20 mL of DMF; and then 5.85 mL (0.0336 mol) ofN,N-diisopropylethylamine were added. The resulting mixture was stirredat room temperature under nitrogen for 6-8 hours while maintaining thetemperature at 20-25° C. Methanol (95 mL) was added in one portion andthen 5.2 mL (0.0672) of trifluoroacetic acid were added within 1 minute.The mixture was stirred for 0.25 hours and then 1.39 g (0.016 mol) ofborane-tert-butyl amine complex were added to the reaction mixture inone portion. The addition funnel was rinsed with 5 mL of methanol, andthe resulting mixture was stirred for 2 hours at room temperature.tert-Butylamine (10.6 mL, 0.101 mol) was added in one portion and theresulting mixture was stirred at 40-42° C. for about 7 hours. Thereaction mixture was then cooled to room temperature and 140 mL of 0.5 NHCl were added, followed by 600 mL of a 10% brine solution at roomtemperature. The resulting mixture was stirred for 2 hours at 20-25° C.,and then cooled to 10° C. and stirred for 1 hour. The resultingprecipitate is collected using a 12.5 cm Buchner funnel by filtering thereaction mixture over a period of about 90 min. The wet cake was washedwith cold water (2×50 mL) and sucked dry for 5 hours. The resultingmaterial was added to 200 mL of acetonitrile while stirring to 2 hoursat 20-25° C. The resulting slurry was filtered through an 8 cm Buchnerfunnel and the collected wet cake was washed with acetonitrile (2×25 mL)and dried under house vacuum (about 25 mm Hg) for 13 hours to afford31.1 g of crude N^(van)-2-(n-decylamino)ethyl vancomycin hydrochloride.This intermediate may then be used in step (c) of the process asdescribed in Example 3.

Example 3 Preparation of N^(van)-2-(n-Decylamino)ethyl29-{[(Phosphonomethyl)amino]methyl}Vancomycin

A 250 mL of three-necked round bottom flask equipped with a mechanicalstirrer, a thermometer and a nitrogen outlet was charged with 5 g ofN^(van)-2-(n-decylamino)ethyl vancomycin and 1.6 g ofaminomethylphosphonic acid and 30 mL of acetonitrile. The slurry wasstirred for 15 minutes to allow disperse solids at 20-30° C. and then 20mL of water was added. The mixture was agitated for 15 minutes and 7.5 gof diisopropylethylamine was added. The resulting mixture was agitateduntil all solids dissolved. The reaction mixture was then cooled to −5to −10° C. and 2.5 g of 3.7% aqueous formaldehyde was charged and theresulting mixture was agitated at −5 to −10° C. for 24 hours. Thereaction was monitored by HPLC. After the reaction was complete, thereaction mixture was adjusted to pH 2-3 with 3M hydrochloric acidsolution while maintaining the reaction temperature at −10 to 5° C. Withmoderate agitation, 125 mL of acetonitrile was added to the reactionmixture at 20 to 25° C. over 10 minutes. The resulting mixture wasstirred at 20 to 25° C. for 2 hours and then filtered. The wet cake waswashed with 20 mL of acetonitrile twice and dried for 18 hours in avacuum oven at 20 to 25° C. to give 5.3 g of the title compound as amixture of the di- and trihydrochloride salt in ˜100% yield with apurity of ca. 80% (HPLC area) (i.e., a compound of formula I where R¹ is—CH₂CH₂—, R² is n-decyl and R⁴ is —CH₂—).

Example 4 Preparation of N^(van)-2-(n-Decylamino)ethyl29-{[(Phosphonomethyl)amino]methyl}Vancomycin

To a 12-L jacketed three-necked flask equipped with a mechanicalstirrer, nitrogen inlet and temperature probe was added 117 g (ca. 60mmol) of N^(van)-2-(n-decylamino)ethyl vancomycin (ca. 80% a purity).Aminomethylphosphonic acid (30 g, 320 mmol) was then added, followed by420 mL of acetonitrile. The resulting slurry was stirred for 15 minutesand then 426 g of water was added and stirring continued for 15 minutes.Diisopropylethylamine (144 g, 1500 mmol) was added ant the mixture wasstirred at room temperature for 1 hour. The resulting light pinksolution was cooled to −7° C. (internal temperature) and 4.51 g (60mmol) of 37% aqueous formaldehyde in 33 mL of acetonitrile were added.The resulting mixture was stirred at −7° C. (internal temperature) for12 hours while monitoring the reaction by HPLC. After the reaction wascomplete (i.e., <1% starting material after 12 hours), the pH of thereaction mixture was adjusted from 10.4 to 2.59 by addition of 3 Naqueous hydrochloric acid solution while maintaining the internalreaction temperature at −4 to −5° C. The amount of 3 N aqueoushydrochloride acid used was 455 g. To the resulting mixture was added3.1 kg of 95% ethanol at 5° C. and the mixture was stirred for 3 hours,and then filtered through a Buchner funnel. The resulting wet cake waswashed with 500 g of ethyl acetate to give 135 g of a granular solid.This solid was dried at 30 mmHg at room temperature for 20 hours to give116 g of the title compound as a mixture of the di- and trihydrochloridesalt. Karl Fisher assay of this material showed an 11% water content;and HPLC analysis showed 1.7% unreacted glycopeptide and 3.6%bis-Mannich byproduct relative to the title compound.

While the present invention has been described with reference to thespecific embodiments thereof, it should be understood by those skilledin the art that various changes may be made and equivalents may besubstituted without departing from the true spirit and scope of theinvention. In addition, many modifications may be made to adapt aparticular situation, material, composition of matter, process, processstep or steps, to the objective, spirit and scope of the presentinvention. All such modifications are intended to be within the scope ofthe claims appended hereto. Additionally, all publications, patents, andpatent documents cited hereinabove are incorporated by reference hereinin full, as though individually incorporated by reference.

1. A process for preparing a compound of formula I:

wherein R¹ is selected from the group consisting of C₁₋₁₀ alkylene,C₂₋₁₀ alkenylene and C₂₋₁₀ alkynylene; R² is selected from the groupconsisting of C₁₋₂₀ alkyl, C₂₋₂₀ alkenyl, C₂₋₂₀ alkynyl, C₃₋₈cycloalkyl, C₅₋₈ cycloalkenyl, C₆₋₁₀ aryl, C₂₋₉ heteroaryl, C₂₋₉heterocyclic, —R^(a)-Cy¹, —R^(a)—Ar¹—Ar², —R^(a)—Ar¹—R^(b)—Ar²,—R^(a)—Ar¹—O—R^(b)—Ar²; R⁴ is C₁₋₁₀ alkylene; R^(a) is selected from thegroup consisting of C₁₋₁₀ alkylene, C₁₋₁₀ alkenylene and C₁₋₁₀alkynylene; R^(b) is selected from the group consisting of C₁₋₆alkylene, C₁₋₆ alkenylene and C₁₋₆ alkynylene; Cy¹ is selected from thegroup consisting of C₃₋₈ cycloalkyl, C₅₋₈ cycloalkenyl, C₆₋₁₀ aryl, C₂₋₉heteroaryl, C₂₋₉ heterocyclic; Ar¹ and Ar² are independently selectedfrom C₆₋₁₀ aryl and C₂₋₉ heteroaryl; wherein each aryl, heteroaryl andheterocyclic group is optionally substituted with 1 to 3 substituentsindependently selected from the group consisting of C₁₋₆ alkyl, C₁₋₆alkoxy, halo, hydroxy, nitro and trifluoromethyl, and each heteroaryland heterocyclic group contains from 1 to 3 heteroatoms selected fromnitrogen, oxygen or sulfur; or a salt thereof; the process comprising:(a) reacting vancomycin or a salt thereof, with a compound of formulaII:

wherein R¹ and R² are as defined herein; and R³ is a amine-labileprotecting group; and a reducing agent to form a compound of formulaIII:

wherein R¹, R² and R³ are as defined herein, or a salt thereof; (b)reacting the compound of formula III with an amine to provide a compoundof formula IV:

wherein R¹ and R² are as defined herein, or a salt thereof; wherein step(a) and step (b) are conducted in the same reaction mixture withoutisolation of the intermediate from step (a). (c) reacting the compoundof formula IV with formaldehyde and a compound of formula V:

in the presence of a base to provide a compound of formula I, or a saltthereof
 2. The process according to claim 1, wherein R¹ is C₁₋₆alkylene.
 3. The process according to claim 2, wherein R¹ is —CH₂—. 4.The process according to claim 1, wherein R² is C₆₋₁₄ alkyl.
 5. Theprocess according to claim 4, wherein R² is n-decyl.
 6. The processaccording to claim 1, wherein R³ is a group of formula (A):W—OC(O)—  (A) wherein W is selected from the group consisting of9-fluorenylmethyl, 3-indenylmethyl, benz[f]inden-3-ylmethyl,17-tetrabenzo[a,c,g,i]fluorenylmethyl,2,7-di-tert-butyl[9-(10,10-dioxo-10,10,10,10-tetrahydrothioxanthyl)]methyl,1,1-dioxobenzo[b]thiophene-2-ylmethyl, wherein the 9-fluorenylmethylgroup is optionally substituted with 1 to 3 substitutents selected fromthe group consisting of C₁₋₆ alkyl, halo, nitro and sulfo.
 7. Theprocess according to claim 6, wherein W is 9-fluorenylmethyl, whereinthe 9-fluorenylmethyl group is optionally substituted with 1 to 3substitutents selected from the group consisting of C₁₋₆ alkyl, halo,nitro and sulfo.
 8. The process according to claim 7, wherein W is9-fluorenylmethyl.
 9. The process according to claim 1, wherein R⁴ isC₁₋₆ alkylene.
 10. The process according to claim 9, wherein R⁴ is—CH₂—.
 11. The process according to claim 1, wherein the reducing agentin step (a) is amine/borane complex.
 12. The process according to claim11, wherein the reducing agent in step (a) is pyridine/borane ortert-butylamine/borane.
 13. The process according to claim 1, whereinstep (a) comprises the steps of: (i) combining vancomycin or a saltthereof with a compound of formula II in the presence of base to form areaction mixture; (ii) acidifying the reaction mixture from step (i)with an acid; and (iii) contacting the reaction mixture from step (ii)with a reducing agent.
 14. The process according to claim 13, whereinthe base in step (i) is a tertiary amine.
 15. The process according toclaim 14, wherein the base in step (i) is diisopropylethylamine.
 16. Theprocess according to claim 13, wherein the acid in step (ii) istrifluoroacetic acid or acetic acid.
 17. The process according to claim1, wherein the amine in step (b) is ammonium hydroxide or a primaryamine.
 18. The process according to claim 17, wherein the amine in step(b) is ammonium hydroxide, methylamine or tert-butylamine.
 19. Theprocess according to claim 18, wherein the amine in step (b) istert-butylamine.
 20. The process according to claim 1, wherein the basein step (c) is a tertiary amine.
 21. The process according to claim 20,wherein the base in step (c) is diisopropylethylamine.
 22. The processaccording to claim 20, wherein the molar ratio of tertiary amine tocompound of formula V is about 3:1 to about 5:1
 23. The processaccording to claim 22, wherein the molar ratio is about 4:1.
 24. Theprocess according to claim 1, wherein R¹ is —CH₂—; R² is n-decyl; R³ isW—OC(O)—, where W is 9-fluorenylmethyl; R⁴ is —CH₂—; the reducing agentin step (a) is tert-butylamine/borane; and the amine in step (b) istert-butylamine.